title: Recombinant expression of homodimeric 660 kDa human thyroglobulin in soybean seeds: an alternative source of human thyroglobulin.
abstract: Soybean seeds possess many qualities that make them ideal targets for the production of recombinant proteins. However, one quality often overlooked is their ability to stockpile large amounts of complex storage proteins. Because of this characteristic, we hypothesized that soybean seeds would support recombinant expression of large and complex proteins that are currently difficult or impossible to express using traditional plant and non-plant-based host systems. To test this hypothesis, we transformed soybeans with a synthetic gene encoding human thyroglobulin (hTG)-a 660 kDa homodimeric protein that is widely used in the diagnostic industry for screening and detection of thyroid disease. In the absence of a recombinant system that can produce recombinant hTG, research and diagnostic grade hTG continues to be purified from cadaver and surgically removed thyroid tissue. These less-than-ideal tissue sources lack uniform glycosylation and iodination and therefore introduce variability when purified hTG is used in sensitive ELISA screens. In this study, we report the successful expression of recombinant hTG in soybean seeds. Authenticity of the soy-derived protein was demonstrated using commercial ELISA kits developed specifically for the detection of hTG in patient sera. Western analyses and gel filtration chromatography demonstrated that recombinant hTG and thyroid-purified hTG are biologically similar with respect to size, mass, charge and subunit interaction. The recombinant protein was stable over three generations and accumulated to ~1.5% of total soluble seed protein. These results support our hypothesis that soybeans represent a practical alternative to traditional host systems for the expression of large and complex proteins.
authors.affiliation: Department of Biology, University of North Carolina at Charlotte, 9201 University City Blvd., Charlotte, NC 28223, USA.
authors.forename: Rebecca
authors.initials: R
authors.lastname: Powell
authors.forename: Laura C
authors.initials: LC
authors.lastname: Hudson
authors.forename: Kevin C
authors.initials: KC
authors.lastname: Lambirth
authors.forename: Diane
authors.initials: D
authors.lastname: Luth
authors.forename: Kan
authors.initials: K
authors.lastname: Wang
authors.forename: Kenneth L
authors.initials: KL
authors.lastname: Bost
authors.forename: Kenneth J
authors.initials: KJ
authors.lastname: Piller
mesh_terms: D015153:Blotting, Western; D002850:Chromatography, Gel; D004797:Enzyme-Linked Immunosorbent Assay; D015870:Gene Expression; D005813:Genes, Synthetic; D005822:Genetic Vectors; D006801:Humans; D018613:Microscopy, Confocal; D030821:Plants, Genetically Modified / Q000235:genetics / Q000378:metabolism; D055550:Protein Stability; D011994:Recombinant Proteins / Q000378:metabolism*; D012231:Rhizobium / Q000235:genetics / Q000378:metabolism; D012639:Seeds / Q000235:genetics / Q000378:metabolism*; D013025:Glycine max / Q000235:genetics / Q000378:metabolism*; D013954:Thyroglobulin / Q000235:genetics / Q000378:metabolism*; D014170:Transformation, Genetic; D019076:Transgenes
chemical_list: D011994:Recombinant Proteins; D013954:Thyroglobulin